The fundamental objective of this project is to determine the differentiation program of murine natural killer (NK) and killer (K) cells and to define the modulating effects of neoplasia on the production of NK and K cells. In vivo cytokinetic studies with 3H-TdR, repopulation assays, andin vitro colony-forming assays are being used to determine the proliferative status and renewal rate of NK and K cells in the bone marrow and their life span in the periphery. Since NK cells acquire binding ability prior to lytic ability, we determined the proliferative status of target binding cells (TBC) in marrow. Six-week-old C57BL/6 female mice were injected intravenously with 3H-TdR (1 micro Curie/gm body weight). Femoral marrow was obtained after 2, 12, 24, or 48 hrs and treated with anti-Ig and C to eliminate B cells. To further select for TBC, erythroid and myeloid cells were eliminated by flow cytometry on the basis of forward and right angle light scatter. The remaining "lymphoid" population comprised 17% of the initial BMC. BM "lymphoid" cells were mixed with FITC-labeled YAC-1 cells in a standard target binding assay. Cytospots were radioautographed and subsequently examined with phase contrast fluorescence microscopy to assess the percent TBC. Two size classes of TBC could be distinguished on the basis of cell diameter: large, greater than 9 micrometers, and small, less than 9 micrometers. The percent of 3H-TdR-labeled small TBC in the marrow was 8, 16, 27, and 16% at 2, 12, 24, and 48 hrs, respectively, whereas the percent labeled large TBC was 24, 18, 21, and 14% at the four time points. The increase in labeling index of small TBC during the first 24 hrs and subsequent decrease indicate that small TBC are newly produced cells with a rapid turnover. The large marrow TBC appear to be a self-maintaining precursor pool that gives rise to small TBC. NK-specific allo-antisera, NK 1.1 and NK 2.1, and functional assays are used to probe the developmental and functional heterogeneity of NK and K cells. We are continuing to test the effect of in vivo depletion of NK and K cells during the neonatal period using our model for specific depletion by in vivo treatment with NK\1.1 antiserum. In the future, we will test the effects of specific depletion of NK cells on: (1)\the production and maturation of NK and K cells, and (2)\the induction and growth of 3-methylcholanthrene-\and Moloney sarcoma virus-induced sarcomas. The effects of tumor growth on the production and life span of NK and K cells will also be determined. (SR)